A genome-wide functional assay of signal transduction in living mammalian cells

被引：56

作者：

Whitney, M ^{[1
]}

Rockenstein, E ^{[1
]}

Cantin, G ^{[1
]}

Knapp, T ^{[1
]}

Zlokarnik, G ^{[1
]}

Sanders, P ^{[1
]}

Durick, K ^{[1
]}

Craig, FF ^{[1
]}

Negulescu, PA ^{[1
]}

机构：

[1] Aurora Biosci Corp, San Diego, CA 92121 USA

来源：

NATURE BIOTECHNOLOGY | 1998年 / 16卷 / 13期

关键词：

gene trap; beta-lactamase; flow cytometry; T-cell;

D O I：

10.1038/4302

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

We describe a genome-wide functional assay for rapid isolation of cell clones and genetic elements responsive to specific stimuli. A promoterless p-lactamase reporter gene was transfected into a human T-cell line to generate a living library of reporter-tagged clones. When loaded with a cell-permeable fluorogenic substrate, the cell library simultaneously reports the expression of a large number of endogenous genes. Flow cytometry was used to recover individual clones whose reporter-tagged genes were either induced or repressed following T-ceIl activation. Responsive clones were expanded and analyzed pharmacologically to identify patterns of regulation associated with specific genes. Although demonstrated using T cells, the genomic assay could be applied to map downstream transcriptional consequences for any propagating cell line in response to any stimulus of interest.

引用

收藏

页码：1329 / 1333

页数：5

相关论文

共 28 条

[1] Differential signaling by lymphocyte antigen receptors [J].

AlberolaIla, J ;

Takaki, S ;

Kerner, JD ;

Perlmutter, RM .

ANNUAL REVIEW OF IMMUNOLOGY, 1997, 15 :125-154

[2] Accessing genetic information with high-density DNA arrays [J].

Chee, M ;

Yang, R ;

Hubbell, E ;

Berno, A ;

Huang, XC ;

Stern, D ;

Winkler, J ;

Lockhart, DJ ;

Morris, MS ;

Fodor, SPA .

SCIENCE, 1996, 274 (5287) :610-614

[3] Evidence for the stochastic integration of gene trap vectors into the mouse germline [J].

Chowdhury, K ;

Bonaldo, P ;

Torres, M ;

Stoykova, A ;

Gruss, P .

NUCLEIC ACIDS RESEARCH, 1997, 25 (08) :1531-1536

[4] IDENTIFICATION OF CALCINEURIN AS A KEY SIGNALING ENZYME IN LYMPHOCYTE-T ACTIVATION [J].

CLIPSTONE, NA ;

CRABTREE, GR .

NATURE, 1992, 357 (6380) :695-697

[5] INDUCTION OF INTERLEUKIN-2 MESSENGER-RNA INHIBITED BY CYCLOSPORIN-A [J].

ELLIOTT, JF ;

LIN, YA ;

MIZEL, SB ;

BLEACKLEY, RC ;

HARNISH, DG ;

PAETKAU, V .

SCIENCE, 1984, 226 (4681) :1439-1441

[6]

Faergeman NJ, 1997, BIOCHEM J, V323, P1

[7] SINGLE CELL ASSAY OF A TRANSCRIPTION FACTOR REVEALS A THRESHOLD IN TRANSCRIPTION ACTIVATED BY SIGNALS EMANATING FROM THE T-CELL ANTIGEN RECEPTOR [J].

FIERING, S ;

NORTHROP, JP ;

NOLAN, GP ;

MATTILA, PS ;

CRABTREE, GR ;

HERZENBERG, LA .

GENES & DEVELOPMENT, 1990, 4 (10) :1823-1834

[8] An induction gene trap screen in embryonic stem cells: Identification of genes that respond to retinoic acid in vitro [J].

Forrester, LM ;

Nagy, A ;

Sam, M ;

Watt, A ;

Stevenson, L ;

Bernstein, A ;

Joyner, AL ;

Wurst, W .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1996, 93 (04) :1677-1682

[9] RAPID PRODUCTION OF FULL-LENGTH CDNAS FROM RARE TRANSCRIPTS - AMPLIFICATION USING A SINGLE GENE-SPECIFIC OLIGONUCLEOTIDE PRIMER [J].

FROHMAN, MA ;

DUSH, MK ;

MARTIN, GR .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1988, 85 (23) :8998-9002

[10] Selection for retroviral insertions into regulated genes [J].

Gogos, JA ;

Lowry, W ;

Karayiorgou, M .

JOURNAL OF VIROLOGY, 1997, 71 (02) :1644-1650